Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Mannosidases”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 19 recordsLinked to original sources
PubMed1985

Marked differences in the swainsonine inhibition of rat liver lysosomal alpha-D-mannosidase, rat liver Golgi mannosidase II, and jack bean alpha-D-mannosidase.

Swainsonine, a plant toxin, strongly inhibits certain alpha-D-mannosidases but has no effect on others [D. R. P. Tulsiani, T. M. Harris, and O. Touster (1982) J. Biol. Chem. 257, 7936-7939]. The reversible inhibition of jack bean and lysosomal alpha-D-mannosidases has previously been suggested to be similar in nature but quite complex. Specific differences in the action of swainsonine on these two enzymes and on Golgi mannosidase II are reported. (a) The inhibition of the jack bean mannosidase, but not rat liver lysosomal alpha-D-mannosidase or Golgi mannosidase II, is increased by preincubation with the alkaloid. (b) The inhibition of the jack bean and lysosomal enzymes, but not mannosidase II, is competitive at inhibitor concentrations of less than or equal to 0.5 microM. (c) The inhibition of jack bean alpha-mannosidase is largely irreversible, its very limited reversibility being partially dependent upon the swainsonine concentration used and on the time of preincubation with the inhibitor. On the other hand, the inhibition of lysosomal alpha-mannosidase is largely reversible, as shown by dilution experiments and by the use of [3H]swainsonine. Golgi mannosidase II shows intermediate reversibility, the results indicating two modes of binding; one rapid and irreversible, the other much slower and reversible.

Alkaloids↗
PubMed1982

alpha-D-Mannosidases of rat liver Golgi membranes. Mannosidase II is the GlcNAcMAN5-cleaving enzyme in glycoprotein biosynthesis and mannosidases Ia and IB are the enzymes converting Man9 precursors to Man5 intermediates.

Current evidence indicates that the trimming of mannosyl residues from intermediates in the biosynthesis of the N-linked oligosaccharides of glycoproteins occurs in the Golgi complex. We now present evidence that mannosidase II (Tulsiani, D. R. P., Opheim, D. J., and Touster, O. (1977) J. Biol Chem. 252, 3227-3233) is the Golgi enzyme that converts GlcNAc Man5 species to GlcNAcMan3 species in completing the mannosyl trimming process required in the biosynthesis of complex type glycoproteins. GlcNAc([3H]Man)5GlcNAc-mannosidase and p-nitrophenyl alpha-D-mannosidase activities copurify throughout the preparative procedure and show the same properties. In addition to mannosidase IA (Tabas, I., and Kornfeld, S. (1979) J. Biol. Chem. 254, 11655-11663), a second alpha-1,2-mannosidase (mannosidase IB) can be prepared from Golgi membranes which is effective in converting Man9GlcNAc to Man5GlcNAc. The two alpha-1,2-mannosidases are very similar in catalytic properties, but they are also distinguishable by several criteria. Although these two enzymes have not been extensively purified, several lines of evidence lead to the tentative conclusion that they are distinct enzymes. They appear to be present in comparable activities in the Golgi membranes and together account for the alpha-1,2-mannosidase activity of these membranes. The particular role of each alpha-1,2-mannosidase remains to be determined.

Animals↗
PubMed1983

Swainsonine, a potent mannosidase inhibitor, elevates rat liver and brain lysosomal alpha-D-mannosidase, decreases Golgi alpha-D-mannosidase II, and increases the plasma levels of several acid hydrolases.

Swainsonine, a toxic plant alkaloid reported to be the agent that induces in animals a neurological condition very similar to the hereditary lysosomal storage disease mannosidosis, and to inhibit the formation of complex glycoproteins of the asparagine-linked class, was recently shown [D.R.P. Tulsiani, T.M. Harris, and O. Touster, (1982) J. Biol. Chem. 257, 7936-7939] to be a highly potent and specific inhibitor of Golgi mannosidase II in addition to being a strong inhibitor of lysosomal mannosidase. In the present study the effect of administered swainsonine on tissue enzyme levels was investigated. The activity of Golgi mannosidase II was markedly decreased (22% of control) without changes occurring in the activities of several other Golgi enzymes. However, the effects of swainsonine on lysosomal enzymes was unexpected. In liver, acid mannosidase increased markedly, instead of decreasing as would be expected from a compound reported to induce a mannosidosis-like condition. Similarly, the principal change in brain was a substantial increase in lysosomal mannosidase levels. In plasma, most lysosomal enzymes increased. These results indicate that the pathological effects of swainsonine are not solely attributable to its being an inhibitor of lysosomal alpha-D-mannosidase and are probably a consequence of abnormal processing of glycoproteins.

Alkaloids↗
PubMed1986

The soluble form of rat liver alpha-mannosidase is immunologically related to the endoplasmic reticulum membrane alpha-mannosidase.

The soluble alpha-mannosidase of rat liver, originally described as a cytoplasmic alpha-mannosidase, has been purified to homogeneity by conventional techniques. The purified enzyme has an apparent molecular weight of 350,000 and is composed of 107-kDa subunits. The soluble alpha-mannosidase has the same enzymatic properties as the endoplasmic reticulum (ER) membrane alpha-mannosidase of rat liver (Bischoff, J., and Kornfeld, R. (1983) J. Biol. Chem. 258, 7909-7910) which is believed to play a role in oligosaccharide processing in the rough ER. Like the membrane-bound ER alpha-mannosidase, the soluble alpha-mannosidase can hydrolyze alpha-linked mannose from both p-nitrophenyl alpha-mannoside (Km = 0.14 mM) and high mannose oligosaccharides, is not inhibited by the mannose analogues swainsonine and 1-deoxymannojirimycin, is stabilized by MnCl2 or CoCl2, and does not bind to concanavalin A-Sepharose. A goat polyclonal antibody raised against the purified soluble alpha-mannosidase specifically recognizes the rat liver membrane-bound ER alpha-mannosidase, leading us to propose that they are two forms of the same enzyme and that the soluble form is derived from the ER membrane alpha-mannosidase by proteolysis. The antibody also cross-reacts with both the soluble and membrane-bound forms of ER alpha-mannosidase activity in cultured Chinese hamster ovary cells and rat H35 hepatoma cells. Since the ER alpha-mannosidase is presumed to be involved in the early steps of oligosaccharide processing, the action of the purified soluble form of the enzyme on high mannose oligosaccharides was examined. Surprisingly, the enzyme released free mannose from oligosaccharides ranging in size from Glc1Man9GlcNAc to Man5GlcNAc with almost equal efficiency. However, a long term incubation of the enzyme with Man9GlcNAc led to the accumulation of Man7GlcNAc and produced only small amounts of Man6GlcNAc and Man5GlcNAc. Structural analysis of these reaction products indicated that the purified soluble form of ER alpha-mannosidase shows little specificity for which mannose residues it removes from Man9GlcNAc. In contrast, as shown in the accompanying paper, the intracellular action of ER alpha-mannosidase on glycoprotein-bound Man9GlcNAc2 is highly specific.

Animals↗
PubMed1996

Endoplasmic reticulum kifunensine-resistant alpha-mannosidase is enzymatically and immunologically related to the cytosolic alpha-mannosidase.

Studies were undertaken to evaluate the relationship of the recently described (S. Weng and R. G. Spiro, 1993, J. Biol. chem. 268, 25656-25663) rat liver kifunensine (KIF)-resistant mannosidase (ER mannosidase II) to the mannose-trimming enzyme of cytosol. We observed that the ER mannosidase II manifests a large number of catalytic and immunological properties similar to those of the cytosolic alpha-mannosidase, which contrast with the quite different characteristics of the KIF-sensitive enzyme (ER mannosidase I). In addition to a mutual resistance to KIF inhibition, the cytosolic enzyme and ER mannosidase II have comparable susceptibility to blocking by swainsonine and 1,4-dideoxy-1,4-imino-D-mannitol, and the latter agent was found to function effectively both in vitro and in vivo. The cytosolic and ER II mannosidases were alike in specifically excising the terminal mannose of the alpha 1,6-linked chain of Man9GlcNAc to yield Man8GlcNAc isomer C; in preferentially hydrolyzing polymannose-GlcNAc1 over polymannose-GlcNAc2 substrates; and in cleaving p-nitrophenyl alpha-D-mannoside. An immunological cross-reactivity between cytosolic mannosidase (M(r) 105 kDa) and ER mannosidase II (M(r) 82 kDa), neither of which is N-glycosylated, was established, suggesting that the latter is translocated posttranslationally into the lumen of the ER compartment in which we found it to be present as a soluble protein. Since antibodies directed against a sequence near the C-terminal end of the cytosolic enzyme reacted with ER mannosidase II while those against a sequence close to the N-terminus did not, it is likely that a proteolytic cleavage of the latter segment takes place during or after translocation. The absence in ER mannosidase II of the pronounced cobalt activation of the cytosolic enzyme suggests that the portion of the polypeptide chain removed during the 105- to 82-kDa conversion includes the binding domain for this ion.

Acetylglucosamine↗
PubMed1992

D-mannonolactam amidrazone. A new mannosidase inhibitor that also inhibits the endoplasmic reticulum or cytoplasmic alpha-mannosidase.

The amidrazone of D-mannonolactam (see compound 5, Fig. 1) was synthesized chemically as a mimic of the mannopyranosyl cation and tested as a potential inhibitor of mannosidases. In this study compound 5 is shown to be a more general mannosidase inhibitor than other currently known compounds and exhibits properties not previously observed with any other mannosidase inhibitors. Thus D-mannonolactam amidrazone not only inhibits the Golgi mannosidase I (IC50 = 4 microM) and mannosidase II (IC50 = 90-100 nM), but it is the first inhibitor that has been shown to be a potent inhibitor of the soluble or endoplasmic reticulum alpha-mannosidase (IC50 = 1 microM). This compound also inhibited the aryl-mannosidases regardless of anomeric configuration although it was much more effective on enzymes recognizing alpha-linked mannose, i.e. jack bean and mung bean alpha-mannosidases (IC50 = 400 nM) as compared with fungal beta-mannosidase (IC50 = 150 microM). Mannonoamidrazone was tested in animal cell cultures using influenza virus-infected Madin-Darby canine kidney cells as a model system, and was found to prevent almost completely the formation of complex types of N-linked oligosaccharides with the formation of about equal amounts of Man9(GlcNAc)2 and Man8(GlcNAc)2 structures. Thus D-mannonolactam amidrazone is a potent but broad spectrum mannosidase inhibitor whose structure and properties should provide valuable insight into the design of other useful glycosidase inhibitors.

Animals↗
PubMed2001

N-glycan structure of a short-lived variant of ribophorin I expressed in the MadIA214 glycosylation-defective cell line reveals the role of a mannosidase that is not ER mannosidase I in the process of glycoprotein degradation.

A soluble form of ribophorin I (RI(332)) is rapidly degraded in Hela and Chinese hamster ovary (CHO) cells by a cytosolic proteasomal pathway, and the N-linked glycan present on the protein may play an important role in this process. Specifically, it has been suggested that endoplasmic reticulum (ER) mannosidase I could trigger the targeting of improperly folded glycoproteins to degradation. We used a CHO-derived glycosylation-defective cell line, MadIA214, for investigating the role of mannosidase(s) as a signal for glycoprotein degradation. Glycoproteins in MadIA214 cells carry truncated Glc(1)Man(5)GlcNAc(2) N-glycans. This oligomannoside structure interferes with protein maturation and folding, leading to an alteration of the ER morphology and the detection of high levels of soluble oligomannoside species caused by glycoprotein degradation. An HA-epitope-tagged soluble variant of ribophorin I (RI(332)-3HA) expressed in MadIA214 cells was rapidly degraded, comparable to control cells with the complete Glc(3)Man(9)GlcNAc(2) N-glycan. ER-associated degradation (ERAD) of RI(332)-3HA was also proteasome-mediated in MadIA214 cells, as demonstrated by inhibition of RI(332)-3HA degradation with agents specifically blocking proteasomal activities. Two inhibitors of alpha1,2-mannosidase activity also stabilized RI(332)-3HA in the glycosylation-defective cell line. This is striking, because the major mannosidase activity in the ER is the one of mannosidase I, specific for a mannose alpha1,2-linkage that is absent from the truncated Man(5) structure. Interestingly, though the Man(5) derivative was present in large amounts in the total protein pool, the two major species linked to RI(332)-3HA shortly after synthesis consisted of Glc(1)Man(5 )and Man(4), being replaced by Man(4 )and Man(3) when proteasomal degradation was inhibited. In contrast, the untrimmed intermediate of RI(332)-3HA was detected in mutant cells treated with mannosidase inhibitors. Our results unambiguously demonstrate that an alpha1,2-mannosidase that is not ER mannosidase I is involved in ERAD of RI(332-)3HA in the glycosylation-defective cell line, MadIA214.

Animals↗
PubMed1998

Substrate specificities of recombinant murine Golgi alpha1, 2-mannosidases IA and IB and comparison with endoplasmic reticulum and Golgi processing alpha1,2-mannosidases.

The catalytic domains of murine Golgi alpha1,2-mannosidases IA and IB that are involved in N-glycan processing were expressed as secreted proteins in P.pastoris . Recombinant mannosidases IA and IB both required divalent cations for activity, were inhibited by deoxymannojirimycin and kifunensine, and exhibited similar catalytic constants using Manalpha1,2Manalpha-O-CH3as substrate. Mannosidase IA was purified as a 50 kDa catalytically active soluble fragment and shown to be an inverting glycosidase. Recombinant mannosidases IA and IB were used to cleave Man9GlcNAc and the isomers produced were identified by high performance liquid chromatography and proton-nuclear magnetic resonance spectroscopy. Man9GlcNAc was rapidly cleaved by both enzymes to Man6GlcNAc, followed by a much slower conversion to Man5GlcNAc. The same isomers of Man7GlcNAc and Man6GlcNAc were produced by both enzymes but different isomers of Man8GlcNAc were formed. When Man8GlcNAc (Man8B isomer) was used as substrate, rapid conversion to Man5GlcNAc was observed, and the same oligosaccharide isomer intermediates were formed by both enzymes. These results combined with proton-nuclear magnetic resonance spectroscopy data demonstrate that it is the terminal alpha1, 2-mannose residue missing in the Man8B isomer that is cleaved from Man9GlcNAc at a much slower rate. When rat liver endoplasmic reticulum membrane extracts were incubated with Man9GlcNAc2, Man8GlcNAc2was the major product and Man8B was the major isomer. In contrast, rat liver Golgi membranes rapidly cleaved Man9GlcNAc2to Man6GlcNAc2and more slowly to Man5GlcNAc2. In this case all three isomers of Man8GlcNAc2were formed as intermediates, but a distinctive isomer, Man8A, was predominant. Antiserum to recombinant mannosidase IA immunoprecipitated an enzyme from Golgi extracts with the same specificity as recombinant mannosidase IA. These immunodepleted membranes were enriched in a Man9GlcNAc2to Man8GlcNAc2-cleaving activity forming predominantly the Man8B isomer. These results suggest that mannosidases IA and IB in Golgi membranes prefer the Man8B isomer generated by a complementary mannosidase that removes a single mannose from Man9GlcNAc2.

Animals↗
PubMed1993

Purification and characterization of rat epididymal-fluid alpha-D-mannosidase: similarities to sperm plasma-membrane alpha-D-mannosidase.

We have previously reported the occurrence and partial characterization of a novel alpha-D-mannosidase activity on rat sperm plasma membranes [Tulsiani, Skudlarek and Orgebin-Crist (1989) J. Cell Biol. 109, 1257-1267]. Here, we report the presence of a similar alpha-D-mannosidase activity in a soluble form in rat epididymal fluid. The soluble enzyme was purified nearly 500-fold with 9-12% recovery to a state approaching homogeneity using: (1) (NH4)2SO4 precipitation; (2) affinity chromatography on immobilized mannan and D-mannosamine; (3) ion-exchange (DE-52) column chromatography; (4) molecular-sieve chromatography. The enzyme was eluted from the final column (Sephacryl S-400) at an apparent molecular mass of 460 kDa. When resolved by SDS/PAGE (under denaturing conditions), the enzyme showed a major protein band (115 kDa) and few very minor bands. The polyclonal antibody raised against the major protein band was found to cross-react with the alpha-D-mannosidase activity present in epididymal fluid (soluble) and detergent-solubilized spermatozoa from the rat and mouse. This result suggested that the soluble and membrane-bound enzyme activities shared a common antigenic site(s). The antibody was used to characterize further the alpha-D-mannosidase activity(ies) present in the rat epididymal fluid and rat sperm plasma membranes. Data from these studies show that the two forms are similar in (a) subunit molecular mass, (b) substrate specificity and (c) inhibitory effect of several sugars. These similarities suggest that the soluble and membrane-bound alpha-D-mannosidase activities are isoforms. Immunoprecipitation studies after solubilization of the testis and epididymal particulate fraction from sexually immature rats show that the testis (but not the epididymis) contains the immunoreactive alpha-D-mannosidase activity. This result and the fact that spermatozoa from the rat rete testis show alpha-D-mannosidase activity indicate that the sperm enzyme is synthesized in the testis during spermatogenesis.

Animals↗
PubMed2000

beta-hexosaminidase, alpha-D-mannosidase, and beta-mannosidase expression in serum from patients with carbohydrate-deficient glycoprotein syndrome type I.

The activity of beta-hexosaminidase, determined with 4-methylumbelliferyl-beta-N-acetylglucopyranoside substrate, and of beta-D-mannosidase was significantly higher in the serum of patients with carbohydrate-deficient glycoprotein (CDG) syndrome type IA (phosphomannomutase deficiency) than in controls. No significant differences were observed in the activity of beta-hexosaminidase, determined using 4-methylumbelliferyl-beta-N-acetylglucopyranoside-6-sulphate as substrate, and the activity of alpha-D-mannosidase. Using DEAE-cellulose chromatography, a greater amount of hexosaminidase B than hexosaminidase A was detected in CDG serum. In CDG serum, hexosaminidase A was eluted in a more basic position in the salt gradient. An isoenzyme of alpha-D-mannosidase and beta-D-mannosidase was identified in control and CDG sera. alpha-D-Mannosidase isoenzyme was eluted in a slightly more basic position in CDG serum than in control serum, whereas beta-D-mannosidase isoenzyme was eluted in the same position.

Adolescent↗
PubMed2005

Regulation of substrate specificity of plant alpha-mannosidase by cobalt ion: in vitro hydrolysis of high-mannose type N-glycans by Co2+-activated Ginkgo alpha-mannosidase.

In our previous study (Woo, K. K., et al., Biosci. Biotechnol. Biochem., 68, 2547-2556 (2004), we purified an alpha-mannosidase from Ginkgo biloba seeds; it was activated by cobalt ions and highly active towards high-mannose type free N-glycans occurring in plant cells. In the present study, we have found that the substrate specificity of Ginkgo alpha-mannosidase is significantly regulated by cobalt ions. When pyridylamino derivative of Man9GlcNAc2 (M9A) was incubated with Ginkgo alpha-mannosidase in the absence of cobalt ions, Man5GlcNAc2-PA (M5A) having no alpha1-2 mannosyl residue was obtained as a major product. On the other hand, when Man9GlcNAc2-PA was incubated with alpha-mannosidase in the presence of Co2+ (1 mM), Man3-1GlcNAc2-PA were obtained as major products releasing alpha1-3/6 mannosyl residues in addition to alpha1-2 mannosyl residues. The structures of the products (Man8-5GlcNAc2-PA) derived from M9A by enzyme digestion in the absence of cobalt ions were the same as those in the presence of cobalt ions. These results clearly suggest that the trimming pathway from M9A to M5A is not affected by the addition of cobalt ions, but that hydrolytic activity towards alpha1-3/6 mannosyl linkages is stimulated by Co2+. Structural analysis of the products also showed clearly that Ginkgo alpha-mannosidase can produce truncated high-mannose type N-glycans, found in developing or growing plant cells, suggesting that alpha-mannosidase might be involved in the degradation of high-mannose type free N-glycans.

Cobalt↗
PubMed2004

Structure of mouse Golgi alpha-mannosidase IA reveals the molecular basis for substrate specificity among class 1 (family 47 glycosylhydrolase) alpha1,2-mannosidases.

Three subfamilies of mammalian Class 1 processing alpha1,2-mannosidases (family 47 glycosidases) play critical roles in the maturation of Asn-linked glycoproteins in the endoplasmic reticulum (ER) and Golgi complex as well as influencing the timing and recognition for disposal of terminally unfolded proteins by ER-associated degradation. In an effort to define the structural basis for substrate recognition among Class 1 mannosidases, we have crystallized murine Golgi mannosidase IA (space group P2(1)2(1)2(1)), and the structure was solved to 1.5-A resolution by molecular replacement. The enzyme assumes an (alphaalpha)(7) barrel structure with a Ca(2+) ion coordinated at the base of the barrel similar to other Class 1 mannosidases. Critical residues within the barrel structure that coordinate the Ca(2+) ion or presumably bind and catalyze the hydrolysis of the glycone are also highly conserved. A Man(6)GlcNAc(2) oligosaccharide attached to Asn(515) in the murine enzyme was found to extend into the active site of an adjoining protein unit in the crystal lattice in a presumed enzyme-product complex. In contrast to an analogous complex previously isolated for Saccharomyces cerevisiae ER mannosidase I, the oligosaccharide in the active site of the murine Golgi enzyme assumes a different conformation to present an alternate oligosaccharide branch into the active site pocket. A comparison of the observed protein-carbohydrate interactions for the murine Golgi enzyme with the binding cleft topologies of the other family 47 glycosidases provides a framework for understanding the structural basis for substrate recognition among this class of enzymes.

Amino Acid Sequence↗
PubMed1992

Human chorionic beta-mannosidase: comparison with beta-mannosidase from human cultured fibroblasts.

The conditions for assay of beta-mannosidase activity in human chorionic villi (CV) were studied using the fluorogenic substrate 4-methylumbelliferyl-beta-D-mannopyranoside. A comparison of the biochemical properties of the CV beta-mannosidase with those of the enzyme from human cultured fibroblasts showed their similarity. Like the enzyme from skin fibroblasts, the CV beta-mannosidase had rather high activity. Both enzymes had virtually the same pH optimum (4.2-4.7) and Km value. The data presented suggest that chorion biopsy specimens can be used for prenatal determination of beta-mannosidase activity at an early stage of development.

Cells, Cultured↗
PubMed1996

Expression of human lysosomal alpha-mannosidase activity in transfected murine cells and human alpha-mannosidase deficient fibroblasts.

We studied the human lysosomal alpha-mannosidase (MANB) by expressing the putative cDNA in mammalian cells, using the eucaryotic expression vector pCDE. The construct pCDE-MANB and pSV2-Neo were cotransfected into human alpha-mannosidase deficient fibroblasts and into a murine cell line and selected by culture in the presence of G418. Six G418 resistant 3T3 clones had increased alpha-mannosidase activity 2 to 3 times above the controls. Two clones from transfected human fibroblasts showed a 2 fold increase in enzyme activity. The human MANB cDNA gene was demonstrated in the target cells by Southern blot analysis and the expression of the gene was shown by RT-PCR analysis. This study is the first to successfully express the MANB gene in a human and a murine cell line. The results confirm that the putative MANB cDNA encodes the full length of lysosomal alpha-mannosidase. Molecular characterization of mannosidosis and approaches to gene therapy are now possible using this cDNA.

3T3 Cells↗
PubMed1978

Lysosomal alpha-D-mannosidase of rat liver. Purification and comparison with the golgi and cytosolic alpha-D-mannosidases.

Rat liver contains alpha-D-mannosidases in lysosomes, Golgi membranes, and cytosol. The lysosomal enzyme has now been purified approximately 30,000-fold over the crude extract and is free of at least 13 other lysosomal hydrolases. The enzyme has an apparent molecular weight of 335,000 by molecular sieve chromatography and 200,000 by sucrose density centrifugation. It is a glycoprotein, as evidenced by its binding to a concanavalin A affinity column and by a positive periodic acid-Schiff stain. The enzyme has a pH optimum near 4.6. Although it is generally insensitive to a large variety of inorganic salts, chelating agents, and sulfhydryl reagents, prolonged exposure to ethylenediaminetetraacetic acid caused loss of activity, which could be restored by the addition of ZnSO4. Substrate specificity studies were performed on the purified lysosomal alpha-D-mannosidase, as well as on the purified Golgi and cytosolic alpha-D-mannosidases. The three enzymes exhibited only very limited activity on native glycoproteins, but were found to be active on glycopeptides and oligosaccharides, hydrolyzing 1 yields 2 and 1 yields 3 linkages, except that the Golgi enzyme had negligible activity towards the latter linkage. Immunological comparisons by antibody precipitation tests and double-diffusion plates indicated that the three enzymes are not immunologically related. The alpha-D-mannosidase isolated from rat epididymis was found to be immunologically very similar, if not identical, to the lysosomal enzyme isolated from rat liver.

Animals↗
PubMed1978

alpha-Mannosidases of genera Aspergillus and Rhizopus. Activity and capacity to utilize Saccharomyces cerevisiae mannan of the best alpha-mannosidase producer Aspergillus flavus Link 69.

Strains of fungi imperfecti of genera Aspergillus and Rhizopus were tested for the ability to produce alpha-mannosidases. The most suitable alpha-mannosidase producer of a total of 20 strains under study was Aspergillus Ravus Link 69. The parameters studied during the cultivation included the growth rate expressed as cell dry weight, alpha-mannosidase activity of the extracellular medium with p-nitorphenyl alpha-D-mannopyranoside as substrate, and utilization of Saccharomyces cerebisiae mannan via its disappearance from the cultivation medium.

Aspergillus↗
PubMed1999

Purification, crystallization and preliminary X-ray crystallographic analysis of recombinant murine Golgi mannosidase IA, a class I alpha-mannosidase involved in Asn-linked oligosaccharide maturation.

Golgi mannosidase IA is a class I alpha-mannosidase which catalyzes the conversion of Man9GlcNAc2 or Man8GlcNAc2 oligosaccharide substrates to Man5GlcNAc2 during the maturation of Asn-linked oligosaccharides. The enzyme is a type II membrane protein, and a recombinant form of mannosidase IA from mouse, lacking the transmembrane domain, has been expressed in Pichia pastoris, purified to homogeneity and crystallized by the hanging-drop vapor-diffusion method. The crystals grow as thin rods, with unit-cell dimensions a = 54.9, b = 135.01, c = 69.9 A. The crystals exhibit the symmetry of space group P2221 and diffract to 2.8 A resolution. The asymmetric unit contains one monomer ( approximately 53 kDa) and has a solvent content of 59%.

Animals↗
PubMed1996

Structure and transcriptional regulation of human alpha-mannosidase IIX (alpha-mannosidase II isotype) gene.

Golgi alpha-mannosidase II is a key enzyme of N-glycan processing. Its genetic defect is associated with HEMPAS (hereditary erythroblastic multinuclearity with positive acidified serum lysis test). We previously cloned cDNAs of human alpha-mannosidase II (alpha-MII) and its isotype, alpha-mannosidase IIX [alpha-MIIX, Misago, M., Liao, Y. F., Eto, S., Mattei. M. G., Moremen. K. W. & Fukuda, M. N. (1995) Proc. Natl Acad. Sci. USA 92, 11766-11770]. Constitutive expressions of alpha-MII and alpha-MIIX mRNA were shown in various human tissues. To investigate the transcriptional regulation of alpha-MIIX gene, we characterized the cosmid clone of 40-kb that includes the 5'-flanking sequence. This clone contains at least eight exons which encode 396 amino acid residues of a total of 1139 amino acid residues of alpha-MIIX. Primer-extension analysis revealed multiple transcription-initiation sites in the range from -70 to -58 relative to the translation-initiation site. No canonical TATA or CAAT boxes were observed, but a (G + C)-rich region was found in close proximity to the transcription-initiation site. To localize the transcriptional regulatory region of this gene, various regions of the 5' sequences were fused to the luciferase gene, and transient-expression assays were conducted in human melanoma G-361 cells. These studies indicated that sequence from -12 to + 11 relative to the most distal 5'-transcription-initiation site was involved in the promoter function. Within this region, the sequence GGGCGT similar to the consensus sequence of the Sp1 binding site, is present at positions -12 to -7. Enhancer activities were found in the region upstream of this site, notably from -4300 to -252. Thus, the alpha-MIIX promoter located in a CpG island is also regulated by upstream elements, indicating the complexity of alpha-MIIX gene expression.

Base Composition↗